Immunoassay‐based Techniques for Early and Specific Detection of Latent Postharvest Anthracnose in Mango

The postharvest anthracnose pathogen Colletotrichum gloeosporioides inciting latent or quiescent infection of mango was detected in early stages using immunoassay methods. Twenty‐five pathotypes isolated from different agroclimatic zones of Tamil Nadu, Karnataka and Pondicherry, India, revealed the variation in protein profile analysis (SDS‐PAGE). The polyclonal antibodies (PCA) were raised against the unfractioned mycelial protein (UMP) and a 40‐kDa polypeptide present in all pathotypes. Standardization of antigen and antiserum dilutions revealed that an antigen dilution of 1 : 200 (protein concentration of 20 μg/ml) and antiserum dilution of 1 : 100 (protein concentration of 40 μg/ml raised against UMP) and 1 : 200 (protein concentration of 20 μg/ml raised against 40 kDa polypeptide) was found to be optimum for the detection of anthracnose pathogen. Both antisera detected the C. gloeosporioides antigen in enzyme‐linked immunosorbent assays (ELISAs), dot immunobinding assays (DIBAs) and Western blots. The specificity in reaction was compared by isolating other Colletotrichum spp. from various hosts viz., C. lindemuthianum (beans), C. falcatum (sugarcane), C. musae (banana), C. capsici (chillies) and Botryodiplodia theobromae (mango). The antisera generated against UMP revealed the cross‐reaction with other host isolates and mango stem end rot pathogen (B. theobromae). The PCA raised against 40‐kDa polypeptide exhibited the specific reaction with C. gloeosporioides isolates in all the immunoassay techniques. By utilizing both PCA, the presence of latent infection was observed in healthy‐looking leaves, flowers and fruits in orchard conditions. The fruit tissues recorded high absorbance values followed by flowers and leaves in all the detection methods. The ELISA technique was also useful in assessing the pathogen inoculum at various biocontrol formulations sprayed mango trees under field conditions. The fluorescent pseudomonad strains mixture (KFP1 + FP7) amended with chitin sprayed at 30‐day intervals revealed the significant reduction in pathogen load than other formulations and unsprayed control.     

<Emphasis Type="Italic">In vitro</Emphasis> plant regeneration via somatic embryogenesis through cell suspension cultures of horsegram [<Emphasis Type="Italic">Macrotyloma uniflorum</Emphasis> (Lam.) verdc.]

Summary In vitro regeneration of plants via somatic embryogenesis through cell suspension culture was achieved in horsegram. Embryogenic calluses were induced on leaf segments on solid Murashige and Skoog (MS) medium with 9.0 μM 2,4-dichlorophenoxyacetic acid (2,4-D). Differentiation of somatic embryos occurred when the embryogenic calluses were transferred to liquid MS medium containing 2,4-D. Maximum frequency (33.2%) of somatic embryos was observed on MS medium supplemented with 7.9 μM 2,4-D. Cotyledonary-torpedo-shaped embryos were transferred to liquid MS medium without growth regulators for maturation and germination. About 5% of the embryos germinated into plants, which grew further on solid MS medium. The plants were hardened and established in soil. Effects of various auxins, cytokinins, carbohydrates, amino acids, and other additives on induction and germination of somatic embryos were also studied. A medium supplemented with 7.9 μM 2,4-D, 3.0% sucrose, 40 mg l⁻¹ L-glutamine, and 1.0 μM abscisic acid was effective to achieve a high frequency of somatic embryo induction, maturation, and further development.     

A new microbial consortia containing entomopathogenic fungus,Beauveria bassiana and plant growth promoting rhizobacteria,Pseudomonas fluorescens for simultaneous management of leafminers and collar rot disease in groundnut

The virulence of 20 isolates of Beauveria bassiana (Balsamo) Vuillemin to larvae of the leafminer, Aproaerema modicella, was tested in the laboratory. Leafminer larvae were sprayed with a standard concentration of 1×10⁸ condia/mL of each B. bassiana isolate. All the B. bassiana isolates tested were pathogenic to A. modicella and the mortality varied between 16.7 and 68.9%. Beauveria bassiana isolate B2 was found to be the most virulent followed by isolate B4 which resulted in 59% mortality. Beauveria isolate B2 was selected for dose–response mortality studies with four different doses (1×10², 1×10⁴, 1×10⁶ and 1×10⁸ conidia/mL). Among the various doses tested, 1×10⁸ conidia/mL produced the highest mortality (70.7%). In addition, morphogenesis of the insect pest in all stages, larval, pupal and adult was greatly affected due to fungal infection. Further, B. bassiana isolate B2 and two Pseudomonas fluorescens strains, TDK1 and Pf1 were tested alone and in combination for suppression of groundnut leafminer and collar rot disease and promotion of plant growth and yield both under glasshouse and field conditions. The mixture of B. bassiana and P. fluorescens strains significantly reduced the leafminer and collar rot disease incidences when applied as talc-based formulation through seed, soil and foliar application under glasshouse and field conditions.     

The MADS box gene, FOREVER YOUNG FLOWER, acts as a repressor controlling floral organ senescence and abscission in Arabidopsis

The ectopic expression of a MADS box gene FOREVER YOUNG FLOWER (FYF) caused a significant delay of senescence and a deficiency of abscission in flowers of transgenic Arabidopsis. The defect in floral abscission was found to be due to a deficiency in the timing of cell separation of the abscission zone cells. Down-regulation of INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) may contribute to the delay of the floral abscission in 35S:FYF flowers. FYF was found to be highly expressed in young flowers prior to pollination and was significantly decreased after pollination, a pattern that correlated with its function. Ethylene insensitivity in senescence/abscission and the down-regulation of ETHYLENE RESPONSE DNA-BINDING FACTOR 1 (EDF1) and EDF2, downstream genes in the ethylene response, in 35S:FYF Arabidopsis suggested a role for FYF in regulating senescence/abscission by suppressing the ethylene response. This role was further supported by the fact that 35S:FYF enhanced the delay of flower senescence/abscission in ethylene response 1 (etr1), ethylene-insensitive 2 (ein2) and constitutive triple response 1 (ctr1) mutants, which have defects in upstream genes of the ethylene signaling pathway. The presence of a repressor domain in the C-terminus of FYF and the enhancement of the delay of senescence/abscission in FYF+SRDX (containing a suppression motif) transgenic plants suggested that FYF acts as a repressor. Indeed, in FYF-DR+VP16 transgenic dominant-negative mutant plants, in which FYF was converted to a potent activator by fusion to a VP16-AD motif, the senescence/abscission of the flower organs was significantly promoted, and the expression of BOP2, IDA and EDF1/2 was up-regulated. Our data suggest a role for FYF in controlling floral senescence/abscission by repressing ethylene responses and regulating the expression of BOP2 and IDA in Arabidopsis.     

Effect of <Emphasis Type="Italic">Tridax procumbens</Emphasis> on liver antioxidant defense system during lipopolysaccharide-induced hepatitis in D-galactosamine sensitised rats

The present study was carried out to assess the effect of chloroform insoluble fraction of ethanolic extract of Tridax procumbens (TP) against D-Galactosamine/Lipopolysaccharide (D-GalN/LPS)-induced hepatitis in rats. Induction of rats with D-GalN/LPS (300 mg/kg body weight/30 g/kg body weight) led to a marked increase in lipid peroxidation as measured by thiobarbituric acid reactive substances (TBARS) in liver. Further there was a decline in the activities of enzymic antioxidants such as superoxide dismutase, catalase, glutathione peroxidase, glutathione s-transferase and the levels of non-enzymic antioxidants namely reduced glutathione, vitamin C and vitamin E. These biochemical alterations were normalised upon pretreatment with TP extract. Thus, the above results suggest that TP (300 mg/kg body weight orally for 10 days) is very effective in allievating the D-GalN/LPS-induced oxidative stress suggesting its antioxidant property. (Mol Cell Biochem 269: 131–136, 2005)     

Effect of Sargassum polycystum (Phaeophyceae)-sulphated polysaccharide extract against acetaminophen-induced hyperlipidemia during toxic hepatitis in experimental rats

The effect of Sargassum polycystum crude extract on lipid metabolism was examined against acetaminophen-induced (800 mg/kg body wt., intraperitoneally) hyperlipidemia during toxic hepatitis in experimental rats. The animals intoxicated with acetaminophen showed significant elevation in the levels of cholesterol, triglycerides and free fatty acid in both serum and liver tissue. The levels of tissue total lipids and serum LDL-cholesterol were also elevated with depleted levels of serum HDL-cholesterol and tissue phospholipid. The acetaminophen-induced animals showed significant alterations in the activities of lipid metabolizing enzymes serum lecithin cholesterol acyl transferase (LCAT) and hepatic triglyceride lipase (HTGL). The levels of liver tissue fatty acids (saturated, mono and polyunsaturated) such as palmitic acid, stearic acid, oleic acid, linoleic acid, arachidonic acid and linolenic acid monitored by gas chromatography were considerably altered in acetaminophen intoxicated animals when compared with control animals. The prior oral administration of Sargassum polycystum (200 mg/kg body wt./day for a period of 15 days) crude extract showed considerable prevention in the severe disturbances of lipid profile and metabolizing enzymes triggered by acetaminophen during hepatic injury. Liver histology also showed convincing supportive evidence regarding their protective nature against fatty changes induced during acetaminophen intoxication. Thus the present study indicates that the protective nature of Sargassum polycystum extract may be due to the presence of active compounds possessing antilipemic property against acetaminophen challenge. (Mol Cell Biochem 276: 89–96, 2005)     

Overexpression of Oncidium MADS box (OMADS1) gene promotes early flowering in transgenic orchid (Oncidium Gower Ramsey)

Oncidium is a popular ornamental orchid and is produced as a high value cash crop for cut flower sold worldwide. Genetically transformed plants of Oncidium were regenerated after cocultivating protocorm-like bodies (PLBs) with Agrobacterium tumefaciens strain LBA4404 harboring pBI121 with OMADS1. The chopped PLBs pre-cultured for 3?days in darkness produced more kanamycin-resistant PLBs. G10 medium containing 200?mg?l^?1 kanamycin was effective for the selection of transformed lines at a frequency of 9%. The rooted plantlets were transferred to pots, acclimated for 3?weeks in the culture room and then moved to the greenhouse. OMADS1 transgene was detected in transgenic lines by PCR, Southern blot analysis and RT-PCR were performed, and the results confirmed that OMADS1 was expressed in these 35S::OMADS1 transgenic plants. CaMV35S::OMADS1 transgenic Oncidium orchid plants flowered significantly earlier, produced more flowers and pseudobulbs than non-transgenic plants. The flower organ conversions were not observed in 35S::OMADS1 transgenic flowers of Oncidium. This is the first report on the ectopic expression of MADS box gene in O. Gower Ramsey using a simple and efficient gene transfer protocol.     

Met receptor tyrosine kinase signaling induces secretion of the angiogenic chemokine interleukin-8/CXCL8 in pancreatic cancer

At diagnosis, the majority of pancreatic cancer patients present with advanced disease when curative resection is no longer feasible and current therapeutic treatments are largely ineffective. An improved understanding of molecular targets for effective intervention of pancreatic cancer is thus urgent. The Met receptor tyrosine kinase is one candidate implicated in pancreatic cancer. Notably, Met is over expressed in up to 80% of invasive pancreatic cancers but not in normal ductal cells correlating with poor overall patient survival and increased recurrence rates following surgical resection. However the functional role of Met signaling in pancreatic cancer remains poorly understood. Here we used RNA interference to directly examine the pathobiological importance of increased Met signaling for pancreatic cancer. We show that Met knockdown in pancreatic tumor cells results in decreased cell survival, cell invasion, and migration on collagen I in vitro. Using an orthotopic model for pancreatic cancer, we provide in vivo evidence that Met knockdown reduced tumor burden correlating with decreased cell survival and tumor angiogenesis, with minimal effect on cell growth. Notably, we report that Met signaling regulates the secretion of the pro-angiogenic chemokine interleukin-8/CXCL8. Our data showing that the interleukin-8 receptors CXCR1 and CXCR2 are not expressed on pancreatic tumor cells, suggests a paracrine mechanism by which Met signaling regulates interleukin-8 secretion to remodel the tumor microenvironment, a novel finding that could have important clinical implications for improving the effectiveness of treatments for pancreatic cancer.